Chronic respiratory infections are a leading cause of death worldwide. These can range in cause from genetic, to toxic, to pathogenic, but nearly all are characterized by dysfunction of the pulmonary immune response. Pathogenic respiratory diseases are caused by airborne bacteria or viruses that are inhaled and withstand the lung immune response to establish infection and cause disease. Of pathogenic bacteria, Mycobacterium tuberculosis (Mtb), the causative agent of Tuberculosis, is by far the deadliest in terms of global burden, infecting nearly a third of the world’s population, and killing 1.5 million per year. Current treatment and prevention strategies of pulmonary tuberculosis are insufficient due to antibiotic resistance and lack of an effective vaccine. Recently, efforts have been made to address this pandemic through the development of host-directed therapies, which, instead of targeting the bacterium directly, aim to treat disease by modulating host immune responses, allowing them to better control the infection. Development of these therapies requires an intimate understanding of the pathogenesis of Mtb within the lungs, the events that take place during infection, and the immune mechanisms that are responsible for a successful or failed immune response. A significant obstacle in developing novel host-directed therapies for pulmonary tuberculosis has been an inability to effectively and efficiently study the tissues that play significant roles during infection. When Mtb enters the lung, the first cell it interacts with is the alveolar macrophage (AM), where it resides for multiple weeks. Therefore, to develop an understanding of the initial events that occur following exposure to Mtb, we must understand the alveolar macrophage and its interactions with Mtb. However, an obstacle in this venture has been the scarcity of these cells within the lungs, and the recalcitrance of these cells to ex vivo culture. In this dissertation, I sought to address this obstacle by developing a novel ex vivo alveolar macrophage model that we can use to expand our understanding of the alveolar macrophage and pulmonary response to disease. We employ CRISPR/Cas9 genetics on a genome-wide scale to identify factors that contribute to the uniqueness of AM biology and pave the way for future genetics approaches to uncover AM-specific mechanisms of pathogen response. We explore the role of the essential cytokine TGFβ in this model and uncover a novel mechanism of TLR2-mediated type 1 interferon production that occurs through a mitochondrial anti-viral response pathway and may play a role in IFNβ production during Mtb infection. To conclude my studies, I investigate a potential genetic interaction of Caspase1 and the phagocyte oxidase during Mtb infection, and discover an extreme susceptibility of mice that lack both of these factors to tuberculosis. This susceptibility is characterized by dysregulation of cytokine responses and recruitment of permissive granulocytes and highlights the utility of the investigation of genetic interactions as a strategy to uncover novel immune mechanisms of protection during tuberculosis. Taken together, these studies explore lung biology and the numerous ways that AMs are a unique and important cell population. They underscore the utility of this platform to, when used in combination with functional genetic approaches, broaden our currently inadequate knowledge of pulmonary tuberculosis and lung inflammation.
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